Determination of the half-life of chloroplast transcripts in tobacco leaves

The amounts of specific transcripts that accumulate in chloroplasts are determined by the rates of synthesis and degradation of the transcripts. The 3' untranslated region of transcripts is a major determinant of the stability of transcripts in chloroplasts. The half-lives of specific transcripts can be determined by northern blot analysis of a time course of transcripts in detached tobacco leaves incubated with actinomycin D, a potent transcription inhibitor. This analysis may be applied to transcripts of endogenous genes or of transgenes introduced into the chloroplast genome in transplastomic plants. Sequence determinants of transcript stability can be identified by analysis of transplastomic plants containing constructs of the green fluorescent protein (gfp) reporter gene fused to the sequences of interest.